Tuesday, September 20, 2011

Microscope Immersion Oil Use

Many microscope manufacturers include a small amount of immersion oil to use with oil immersion objectives when microscopes are sold. Since it only takes a drop to use this material, a small bottle will last for quite a while, but eventually needs replacement.

Lenses designed to be used with immersion oil usually have a black ring around the barrel next to the colored ring identifying the magnification. They will also show “oil” in the identifying markings. These are the only lenses which should be used with oil and they should always be used with oil. The image without the oil interface will show optical aberrations and result in a very poor image.

Air has a refractive index of 1.0 and a good oil lens has a numerical aperture of 1.25 or 1.30. The glass used in slides and coverslips typically has a refractive index of 1.515 The full resolution of the lens can only be achieved by matching the refractive indexes of the materials the light passes through and it will be limited by the lowest number. Using air instead of oil limits the resolution to a lower numerical aperture, less than 1.0, resulting in a lower resolution and less brightness.

The point of an immersion oil objective is a superior image in terms of resolution and brightness, therefore matching the refractive indexes, 1.515 in the glass and the typical immersion oil with a refractive index of 1.515 results in the best possible image. In this case the resolution is improved by approximately 50% over a dry lens with the same magnification.

After use the objective does not have to be cleaned completely if it will be used again within days. However, it is a good idea to blot the front of the lens with lens tissue to remove excess oil. If the microscope will not be used again soon, the oil should be cleaned off. This assumes you will be using the same immersion oil. Anytime you change brands of immersion oil, the lens should be cleaned of any of the previous oil. Mixing oils can result in clouding of the material and a diminished image, not to mention, a new compound with unknown properties.

The biggest mistake made with the use of immersion oil is going back to a lower power objective once oil has been added to the slide. It is a very good habit to examine your sample completely at low power before you oil the slide. If you oil the slide for the 100x microscope objective and go back to the 40x, you will get oil on the 40x and the image will not be sharp. It is much easier to keep oil off the 40x than it is to do a good cleaning after it gets oiled.

Letting oil remain on an objective not designed for oil also increases the potential for the oil to degrade the seal or the lens cement and seep into the interior of the lens assembly. This very effectively ruins the objective which in most cases has to be replaced.